1. Field of the Invention
This invention relates to reaction tubes which are used in performing the Polymerase Chain Reaction (PCR) in thermal cyclers utilizing temperature controlled heat blocks for heating and cooling.
2. Description of the Prior Art
Thin-walled 0.2 ml polypropylene reaction tubes are the standard for performing the Polymerase Chain Reaction (PCR) in thermal cyclers utilizing temperature-controlled heat blocks for heating and cooling, as disclosed, for example, in U.S. Pat. No. 3,725,010 patented Apr. 3, 1973 by Beckman Instruments Inc. Typically, the reaction tube is inserted into a well in the heat block so that the sides are surrounded by the heat block but the cap is not in contact with the heat block. Since the cap is not in contact with the heat block, it is cooler than the sides of the tube. There are a number of solutions that have been invented to minimize evaporation and condensation of liquid reagents into the cap during the heating and cooling cycles of PCR.
These patented solutions include:
U.S. Pat. No. 7,081,600, patented Jul. 25, 2006 by Stratagene California for “Method and Apparatus for Cover Assembly for Thermal Cycling of Samples”. That patent provided a cover assembly for heating a plurality of samples which included a plurality of heater element areas located within a housing, and a force distribution system that engaged the plurality of heater element areas and distributed a force over the plurality of heater element areas, so that the cover assembly flexibly engaged the plurality of samples and heated the plurality of samples. The arrangement of the resistive heater, the heater backing plate, the force distribution system and the support plate allegedly provided substantial temperature uniformity among a plurality of sample tubes for receiving samples of biological material. The flexible heating cover assembly was alleged to improve the uniformity, efficiency, quality, reliability and controllability of the thermal response during thermal cycling of the biological material.
U.S. Pat. No. 6,878,905, patented Apr. 12, 2005 by Stratagene California for “Apparatus and Method For Flexible Heating Cover Assembly For Thermal Cycling of Samples of Biological Material”. The patent provided a flexible heating cover assembly for an apparatus for thermal cycling of samples of biological material which included: a housing including a plurality of assembly skirt components; a resistive heater located within the housing, the resistive heater including at least one outer heater element area and at least one central heater element area; a heater backing plate connected to the resistive heater to protect the resistive heater; a load sharing system engaging the heater backing plate to promote uniform contact of the resistive heater with a plurality of sample tubes; and a support plate aligning the assembly skirt components, so that the flexible heating cover assembly provides non-uniform heat distribution among the samples of biological material. The flexible heating cover assembly was alleged to improve the uniformity, efficiency, quality, reliability and controllability of the thermal response during thermal cycling of the biological material.
U.S. Pat. No. 6,730,883, patented May 4, 2004 by Stratagene California for “Flexible Heating Cover Assembly for Thermal Cycling of Samples of Biological Material”. The patent provided a flexible heating cover assembly which included a plurality of engageable enclosure components; a resistive heater located within the housing, the resistive heater including a plurality of heater element areas; a heater backing plate engaging the resistive heater and providing stability to the resistive heater; a force distribution system that engaged the heater backing plate and distributed a force over the heater backing plate; and a support plate providing stiffness for the force distribution system, so that the arrangement of the resistive heater, the heater backing plate, the force distribution system and the support plate provide substantial temperature uniformity among a plurality of sample tubes for receiving samples of biological material. The arrangement of the resistive heater, the heater backing plate, the force distribution system and the support plate was alleged to provide substantial temperature uniformity among a plurality of sample tubes for receiving samples of biological material. The flexible heating cover assembly was alleged to improve the uniformity, efficiency, quality, reliability and controllability of the thermal response during thermal cycling of the biological material.
U.S. Pat. No. 5,552,580, patented Sep. 3, 1996 by Beckman Instruments Limited for “Heated Cover Device” provided a heated cover for a receptacle containing a vaporizable substance. The cover was heated to a temperature above the temperature of the substance so as to prevent condensation of vapor evaporated from the substance. A device for placing and removing the cover with respect to the receptacle is designed in connection with a temperature-controlled heating/cooling plate which controls the temperature of the contents of the receptacle. Thus, the heated lid heated the cap to a temperature equivalent or hotter than the sides of the
These attempted solutions were not completely effective since, by definition, the use of a heated lid required extra electronics and components compared to a system without a heated lid.
Another proposed solution was to overlay mineral oil atop of the liquid reagents to prevent evaporation at the reaction temperature.
This attempted solution was not completely effective since mineral oil represents an extra reagent that must be added in an extra step. Both of these solutions complicate the process of PCR.
In addition, real-time thermal cyclers combine a thermal cycling apparatus with a system for optical detection of fluorescence generated by a successful PCR. Some Instruments excite and image the reaction through the top of the tube. For example, this is the setup of the “Stepfme” system from Applied Biosystems. That procedure required that the top of the tube be flat and be optically clear.
Other systems excite the reaction from the side and image the reaction from the bottom of the tube. For example, this is the setup of the Spartan DX™ system from Spartan Bioscience. A problem using such system occurs because standard PCR tubes have curved bottoms. The curvature reflects and refracts both the excitation and emission light. This results in light artifacts that reduce sensitivity and specificity of the optical detection.